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Detection method of amyloid protein

A technology for amyloid and samples, applied in measuring devices, instruments, fluorescence/phosphorescence, etc., can solve the problems of interference in the detection process and single detection method, and achieve the effect of good biocompatibility and low cost

Active Publication Date: 2016-06-08
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The method of the present invention is simple, low in cost, low in cost, and the carbon-based quantum dot material has good biocompatibility; the detection process does not interfere with the system; and solves the current technical bottleneck of single detection method and interference introduced in the detection process

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  • Detection method of amyloid protein
  • Detection method of amyloid protein
  • Detection method of amyloid protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Embodiment 1: Preparation of carbon-based quantum dots

[0067] 1) Mix 1g of graphite powder with 200mL of concentrated sulfuric acid and 40g of sodium nitrate, and cool to zero. Subsequently, 4 g of potassium permanganate was slowly added, the temperature was kept at 10° C., and the mixture was vigorously stirred.

[0068] 2) After the above mixture was stirred evenly, it was transferred to an oil bath at 30°C and reacted for 50 minutes. Immediately afterwards, the temperature was raised to 140° C., and the reaction was continued for 20 h.

[0069] 3) After the reaction, the product was cooled to room temperature, diluted with 500 mL of deionized water, and adjusted to about pH 3 with Na2CO3. After filtering through a 0.22 μm membrane filter, take the supernatant. Place in a centrifuge and centrifuge at 10,000 rpm for 10 minutes. Finally, the final quantum dots were obtained by dialysis for 3 days with a dialysis bag with a molecular weight cut-off of 1000 Da.

[...

Embodiment 2

[0072] Example 2: Detection of β-amyloid monomers and fibers by carbon-based quantum dots

[0073] 1) Detection of β-amyloid monomer by carbon-based quantum dots

[0074] Aβ powder was formulated into 1 mg / mL hexafluoroisopropanol (HFIP) solution, incubated at 37°C for 3 hours, blown dry with nitrogen, and dispersed in Six Aβmonomer solutions with concentrations of 50mg / mL, 100mg / mL, 150mg / mL, 200mg / mL, 250mg / mL and 300mg / mL were formed in ultrapure water. Take 100 microliters of each of the above six solutions and mix them with 100 microliters of 1 mg / mL carbon-based quantum dot solution in a 96-well plate. Afterwards, place in a fluorescence spectrometer to measure the fluorescence intensity of the carbon-based quantum dots, set the excitation wavelength to 400nm, and the emission wavelength to 500nm. The curve of the fluorescence intensity of the carbon-based quantum dots changing with the concentration of the Aβ monomer was obtained. curve like Figure 4 As shown in (a)...

Embodiment 3

[0079] Example 3: Detection of carbon-based quantum dots on the content of β-amyloid fibers

[0080] The preparation method of Aβ monomer solution and Aβ fiber solution: same as Example 2.

[0081] 1) Mixing the prepared Aβ monomer and fiber solution according to the mass percentage of Aβ fiber from 0 to 100% to obtain Aβ monomer / fiber mixed solution with different proportions.

[0082] 2) Mix 100 microliters of the above solution with 100 microliters of carbon-based quantum dots (0.8mg / mL), add them into a 96-well plate, set the excitation wavelength to 400nm, and the emission wavelength to 450nm. The fluorescence intensity of carbon-based quantum dots in different proportions of Aβ monomer / fiber mixture was measured. The result is as Figure 5 shown. The results show that the fluorescence intensity of carbon-based quantum dots increases linearly with the increase of fiber content in the mixture.

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Abstract

The invention relates to a detection method of amyloid protein. Specifically, the detection method is realized by means of the unique structure and the luminescent characteristic of carbon-based quantum dots serving as fluorescent probes. A novel scheme is provided for aggregation detection of amyloid protein, and interference to a to-be-detected system in a traditional fluorescent detection method is avoided.

Description

technical field [0001] The invention relates to a method for detecting amyloid, in particular to a method for detecting amyloid by using carbon-based quantum dots, and relates to the field of protein structure detection. Background technique [0002] The occurrence of amyloidosis originates from the misfolding of proteins, which are deposited in body tissues or organs in the form of precipitates, and directly lead to Alzheimer's disease, Parkinson's syndrome, mad cow disease and other diseases (Cell, 2012, 148, 1188.) . The specific aggregation process of amyloid protein is as follows. The protein in the solution is misfolded and transformed into a β-sheet structure, and then assembled into very small oligomers and diffuse small oligomers, circular fibrils, and fibrils with physiological toxicity. , fibers, and eventually form plaques. For example, detecting and tracking the changes of amyloid structure is a research hotspot today. At present, Thioflavin T and Congo red ar...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
CPCG01N21/64G01N21/6486G01N2021/6417
Inventor 杨延莲黄欢李平王琛
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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